The arterivirus porcine reproductive and respiratory syndrome virus (PRRSV) causes significant economic losses to the swine industry worldwide. Here we apply ribosome profiling (RiboSeq) and parallel ...RNA sequencing (RNASeq) to characterise the transcriptome and translatome of both species of PRRSV and to analyse the host response to infection. We calculated programmed ribosomal frameshift (PRF) efficiency at both sites on the viral genome. This revealed the nsp2 PRF site as the second known example where temporally regulated frameshifting occurs, with increasing -2 PRF efficiency likely facilitated by accumulation of the PRF-stimulatory viral protein, nsp1β. Surprisingly, we find that PRF efficiency at the canonical ORF1ab frameshift site also increases over time, in contradiction of the common assumption that RNA structure-directed frameshift sites operate at a fixed efficiency. This has potential implications for the numerous other viruses with canonical PRF sites. Furthermore, we discovered several highly translated additional viral ORFs, the translation of which may be facilitated by multiple novel viral transcripts. For example, we found a highly expressed 125-codon ORF overlapping nsp12, which is likely translated from novel subgenomic RNA transcripts that overlap the 3' end of ORF1b. Similar transcripts were discovered for both PRRSV-1 and PRRSV-2, suggesting a potential conserved mechanism for temporally regulating expression of the 3'-proximal region of ORF1b. We also identified a highly translated, short upstream ORF in the 5' UTR, the presence of which is highly conserved amongst PRRSV-2 isolates. These findings reveal hidden complexity in the gene expression programmes of these important nidoviruses.
Abnormal muscle tone is a common motor disorder following stroke, which may require rehabilitation. The Modified Ashworth Scale is a 6-point rating scale that is used to measure muscle tone. The ...interrater and intrarater reliability of measurements obtained with the scale remain equivocal. The purpose of this study was to investigate the reliability of measurements obtained with the scale in the lower limb of patients with stroke.
Twenty patients were tested 2 weeks after their stroke, and 12 patients were tested 12 weeks after their stroke.
Gastrocnemius, soleus, and quadriceps femoris muscles on the hemiplegic side were tested.
Interrater reliability for 2 raters was poor, with a Kendall tau-b correlation for the combined muscle group of.062 (P=.461). For intrarater reliability, the Kendall tau-b correlation was.567 (P<.001). The agreement within one rater occurred mostly on the grade of 0.
The Modified Ashworth Scale yielded reliable measurements in the lower limb for a single examiner, and agreement was best on the grade of 0. The reliability between examiners was not good, which may bring into question the validity of measurements obtained with the scale.
Abstract
Background
RNA editing at the Q/R site of GluA2 occurs with ~99% efficiency in the healthy brain, so that the majority of AMPARs contain GluA2(R) instead of the exonically encoded GluA2(Q). ...Reduced Q/R site editing increases AMPA receptor calcium permeability and leads to dendritic spine loss, neurodegeneration, seizures and learning impairments. Furthermore, GluA2 Q/R site editing is impaired in Alzheimer’s disease (AD), raising the possibility that unedited GluA2(Q)-containing AMPARs contribute to synapse loss and neurodegeneration in AD. If true, then inhibiting expression of unedited GluA2(Q), while maintaining expression of GluA2(R), may be a novel strategy of preventing synapse loss and neurodegeneration in AD.
Methods
We engineered mice with the ‘edited’ arginine codon (CGG) in place of the unedited glutamine codon (CAG) at position 607 of the
Gria2
gene. We crossbred this line with the J20 mouse model of AD and conducted anatomical, electrophysiological and behavioural assays to determine the impact of eliminating unedited GluA2(Q) expression on AD-related phenotypes.
Results
Eliminating unedited GluA2(Q) expression in AD mice prevented dendritic spine loss and hippocampal CA1 neurodegeneration as well as improved working and reference memory in the radial arm maze. These phenotypes were improved independently of Aβ pathology and ongoing seizure susceptibility. Surprisingly, our data also revealed increased spine density in non-AD mice with exonically encoded GluA2(R) as compared to their wild-type littermates, suggesting an unexpected and previously unknown role for unedited GluA2(Q) in regulating dendritic spines.
Conclusion
The Q/R editing site of the AMPA receptor subunit GluA2 may act as an epigenetic switch that regulates dendritic spines, neurodegeneration and memory deficits in AD.
Neuronal hyperexcitability in Alzheimer’s disease (AD) models is thought to either contribute to the formation of amyloid beta plaques or result from their formation. Neuronal hyperexcitability has ...been shown in the cerebral cortex of the widely used young APPswe/PS1dE9 mice, which have accelerated plaque formation. However, it is currently unclear if hyperexcitability also occurs in CA1 hippocampal neurons of aged animals in this model. In the present work, we have compared intrinsic excitability and spontaneous synaptic inputs from CA1 pyramidal cells of 8-month-old APPswe/PS1dE9 and wildtype control mice. We find no change in intrinsic excitability or spontaneous postsynaptic currents (PSCs) between groups. We did, however, find a reduced input resistance and an increase in hyperpolarization-activated sag current. These results are consistent with findings from other aged AD model mice, including the widely used 5xFAD and 3xTg. Together these results suggest that neuronal hyperexcitability is not a consistent feature of all AD mouse models, particularly at advanced ages.
Objective: To investigate the relative impact of radiographic osteoarthritis (ROA) and current knee pain on lower limb physical function, quadriceps strength, knee joint proprioception, and postural ...sway. Methods: Using a 2×2 factorial design, 142 community derived subjects aged over 45 were divided into four subgroups based on the presence or absence of ROA (Kellgren & Lawrence >grade 2) and knee pain (as assessed by NHANES questions and a 100 mm visual analogue scale). Maximum isometric contraction of the quadriceps, knee joint proprioceptive acuity, static postural sway, and WOMAC index (both whole and function subscale) were assessed in all subjects. Results: Compared with normal subjects, reported disability was greater for all other subgroups (p<0.01). Subjects with both ROA and knee pain reported the greatest disability, and those with knee pain only had greater disability than those with ROA only. Quadriceps weakness was observed in all groups compared with normal subjects (p<0.01), though they were no significant intergroup differences. Subjects with knee pain had a greater sway area than those without (p<0.05) but the presence of ROA was not associated with increased postural sway. No differences in proprioceptive acuity were observed between groups. Conclusions: The presence of knee pain has a negative association with quadriceps strength, postural sway, and disability compared with ROA. However, the presence of pain-free ROA has a significant negative influence on relative quadriceps strength and reported disability.
The Analog Devices TMP37 temperature sensor is used to monitor the differences in temperature between sensors pairs in order to find the gradients in the U.S. Atlas End Cap Muon Chambers. It was ...chosen because of its stability, linearity, high output signal, and especially the low self-heating. We have irradiated samples of these sensors with neutrons and gamma rays. The results of these measurements are presented.
Secreted amyloid precursor protein-alpha (sAPPα) has growth factor-like properties and can modulate long-term potentiation (LTP) and memory. Here, we demonstrate that exposure to sAPPα converts ...short-lasting LTP into protein-synthesis-dependent late LTP in hippocampal slices from male rats. sAPPβ had no discernable effect. We hypothesized that sAPPα facilitated LTP via regulated glutamate receptor trafficking and
protein synthesis. We found using a linear mixed model that sAPPα stimulated trafficking of GluA2-lacking AMPARs, as well as NMDARs to the extrasynaptic cell surface, in a calcium/calmodulin-dependent kinase II and protein kinase G-dependent manner. Both cell surface receptor accumulation and LTP facilitation were present even after sAPPα washout and inhibition of receptor trafficking or protein synthesis prevented all these effects. Direct visualization of newly synthesized proteins (FUNCAT-PLA) confirmed the ability of sAPPα to stimulate
protein synthesis and revealed GluA1 as one of the upregulated proteins. Therefore, sAPPα generates a coordinated synthesis and trafficking of glutamate receptors to the cell surface that facilitate LTP.
Secreted amyloid precursor protein-alpha (sAPPα) is a neurotrophic and neuroprotective protein that can promote synaptic plasticity and memory, yet the molecular mechanisms underlying these effects are still not well understood. Here, we show that sAPPα facilitates long-term potentiation (LTP) in a concentration-dependent fashion through cellular processes involving
protein synthesis and trafficking of both GluA2-lacking AMPARs and NMDARs to the extrasynaptic cell surface. sAPPα also enhances GluA1, but not GluA2, synthesis. The trafficking effects, along with the LTP facilitation, persist after sAPPα washout, revealing a metaplastic capability of exogenous sAPPα administration. sAPPα thus facilitates LTP through coordinated activation of protein synthesis and trafficking of glutamate receptors to the cell surface, where they are positioned for priming LTP.
Alzheimer's disease (AD) is a neurodegenerative disease driven in large part by accumulated deposits in the brain of the amyloid precursor protein (APP) cleavage product amyloid-β peptide (Aβ). ...However, AD is also characterised by reductions in secreted amyloid precursor protein-alpha (sAPPα), an alternative cleavage product of APP. In contrast to the neurotoxicity of accumulated Αβ, sAPPα has many neuroprotective and neurotrophic properties. Increasing sAPPα levels has the potential to serve as a therapeutic treatment that mitigates the effects of Aβ and rescue cognitive function. Here we tested the hypothesis that lentivirus-mediated expression of a human sAPPα construct in a mouse model of AD (APPswe/PS1dE9), begun before the onset of plaque pathology, could prevent later behavioural and electrophysiological deficits. Male mice were given bilateral intra-hippocampal injections at 4 months of age and tested 8-10 months later. Transgenic mice expressing sAPPα performed significantly better than untreated littermates in all aspects of the spatial water maze task. Expression of sAPPα also resulted in partial rescue of long-term potentiation (LTP), tested in vitro. These improvements occurred in the absence of changes in amyloid pathology. Supporting these findings on LTP, lentiviral-mediated expression of sAPPα for 3 months from 10 months of age, or acute sAPPα treatment in hippocampal slices from 18 to 20 months old transgenic mice, completely reversed the deficits in LTP. Together these findings suggest that sAPPα has wide potential to act as either a preventative or restorative therapeutic treatment in AD by mitigating the effects of Aβ toxicity and enhancing cognitive reserve.
Processing of the amyloid precursor protein by alternative secretases results in ectodomain shedding of either secreted amyloid precursor protein-α (sAPPα) or its counterpart secreted amyloid ...precursor protein-β (sAPPβ). Although sAPPα contains only 16 additional amino acids at its C-terminus compared to sAPPβ, it displays significantly greater potency in neuroprotection, neurotrophism and enhancement of long-term potentiation (LTP). In the current study, this 16 amino acid peptide sequence (CTα16) was characterised for its ability to replicate the synaptic plasticity-enhancing properties of sAPPα. An N-acetylated version of CTα16 produced concentration-dependent increases in the induction and persistence of LTP at Schaffer collateral/commissural synapses in area CA1 of young adult rat hippocampal slices. A scrambled peptide had no effect. CTα16 significantly enhanced de novo protein synthesis, and correspondingly its enhancement of LTP was blocked by the protein synthesis inhibitor cycloheximide, as well as by the α7-nicotinic receptor blocker α-bungarotoxin. The impaired LTP of 14–16 month old APPswe/PS1dE9 transgenic mice, a mouse model of Alzheimer's disease, was completely restored to the wild-type level by CTα16. These results indicate that the CTα16 peptide fragment of sAPPα mimics the larger protein's functionality with respect to LTP, stimulation of protein synthesis and activation of α7-nAChRs, and thus like sAPPα may have potential as a therapeutic agent against the plasticity and cognitive deficits observed in AD and other neurological disorders.
•Peptide CTα16 enhanced LTP similarly to secreted amyloid precursor protein-α.•The enhancement of LTP by CTα16 was concentration- and sequence-dependent.•CTα16 stimulated de novo protein synthesis, contributing to enhancement of LTP.•CTα16 completely rescued LTP in a transgenic mouse model of Alzheimer's disease.
Differential processing of the amyloid precursor protein liberates either amyloid-ß, a causative agent of Alzheimer's disease, or secreted amyloid precursor protein-alpha (sAPPα), which promotes ...neuroprotection, neurotrophism, neurogenesis and synaptic plasticity. The underlying molecular mechanisms recruited by sAPPα that underpin these considerable cellular effects are not well elucidated. As these effects are enduring, we hypothesised that regulation of gene expression may be of importance and examined temporally specific gene networks and pathways induced by sAPPα in rat hippocampal organotypic slice cultures. Slices were exposed to 1 nM sAPPα or phosphate buffered saline for 15 min, 2 h or 24 h and sAPPα-associated gene expression profiles were produced for each time-point using Affymetrix Rat Gene 1.0 ST arrays (moderated t-test using Limma: p < 0.05, and fold change ± 1.15).
Treatment of organotypic hippocampal slice cultures with 1 nM sAPPα induced temporally distinct gene expression profiles, including mRNA and microRNA associated with Alzheimer's disease. Having demonstrated that treatment with human recombinant sAPPα was protective against N-methyl d-aspartate-induced toxicity, we next explored the sAPPα-induced gene expression profiles. Ingenuity Pathway Analysis predicted that short-term exposure to sAPPα elicited a multi-level transcriptional response, including upregulation of immediate early gene transcription factors (AP-1, Egr1), modulation of the chromatin environment, and apparent activation of the constitutive transcription factors CREB and NF-κB. Importantly, dynamic regulation of NF-κB appears to be integral to the transcriptional response across all time-points. In contrast, medium and long exposure to sAPPα resulted in an overall downregulation of gene expression. While these results suggest commonality between sAPPα and our previously reported analysis of plasticity-related gene expression, we found little crossover between these datasets. The gene networks formed following medium and long exposure to sAPPα were associated with inflammatory response, apoptosis, neurogenesis and cell survival; functions likely to be the basis of the neuroprotective effects of sAPPα.
Our results demonstrate that sAPPα rapidly and persistently regulates gene expression in rat hippocampus. This regulation is multi-level, temporally specific and is likely to underpin the neuroprotective effects of sAPPα.
PDF
